Introduction of a foreign substance, in the following referred to as antigen (Ag), including hapten, by injection into a vertebrate organism may result in the induction of an immune response characterized by the production of specific antibodies (products of B lymphocytes) capable of interacting with said Ag and/or the development of effector T lymphocytes and the production of soluble mediators, termed lymphokines, at the site of encounter with said Ag. Antibodies and T lymphocytes do certainly play an essential role in protecting against hostile Ag but can also participate in injurious processes leading to destruction of host tissues. This is the case in autoimmune diseases where antibodies and/or T lymphocytes react with Ag of one's own tissues and damage these. This is also the case in allergic reactions characterized by an exaggerated immune response to certain environmental matters and which may result in inflammatory responses leading to tissue destruction. Moreover, this is the case in chronic inflammatory reactions that develop as a result of ineffective elimination of foreign materials as in certain infections (e.g. tuberculosis, schistosomiasis) or following introduction of foreign particles (e.g. asbestos). This is also the case in immunoproliferative reactions that follow the introduction into the body of an allograft and lead to its rejection.
One of the primary goals in developing effective therapies against diseases caused by unwanted or tissue damaging immunological reactions such as allograft rejection, autoimmune diseases, tissue destructive allergic reactions to infectious microorganisms or to environmental antigens, is to specifically suppress or decrease to an acceptable level the intensity of deleterious immune processes without affecting the remainder of the immune system.
The subject of immunological tolerance deals with all mechanisms that ensure an absence ofdestructive immune response, be it to one's own body constituents ("self antigens") or to any given foreign substance.
A long-recognized method of inducing immunological tolerance is the oral administration of antigen which was first demonstrated by Wells for hen egg proteins (Wells, H. 1911. Studies on the chemistry of anaphylaxis III. Experiments with isolated proteins, especially those of hen's egg. J. Infect. Dis. 9:147). The phenomenon, often referred to as "oral tolerance" (because initially documented by the effect of oral administration of Ag), is characterized by the fact that animals fed or having inhaled an antigen become refractory or have diminished capability to develop a systemic immune response when re-exposed to said Ag introduced by the systemic route, e. g. by injection. In broad terms, linking of an antigen onto a mucosal membrane or into a mucosal tissue, be it the intestine, the lung, the mouth, the genital tract, the nose or the eye, can induce the phenomenon of systemic immunological tolerance. As opposed to this, introduction of an antigen into a non mucosal tissue, i.e. for example the skin or the blood, referred to as systemic immunization, often results in an immune response with the characteristics mentioned above, and is referred to as systemic immune response. The phenomenon is highly specific of the Ag introduced by the mucosal route in the sense that hyporesponsiveness can only be documented subsequent to injection of said fed or inhaled Ag but not after injection of a structurally unrelated Ag (provided the latter had not previously been encountered at mucosal sites).
It is believed that ingested antigens are absorbed and processed by specialized cells, including epithelial enterocytes and Peyer's patch M cells, in the gut-associated lymphoid tissue (Owen, R. L., and P. Nemanic. 1978. Antigen processing structures of the mammalian intestinal tract: an SEM study of lymphoepithelial organs. Scanning Electron Microsc. 2: 367-378.). It is also believed that inhaled antigens are taken up by similar types of cells in the airway epithelium (Richardson J, Bouchard R and Ferguson C C. 1976. Uptake and transport of exogenous proteins by respiratory epithelium. Lab. Invest. 35:307-314). Following interaction of the antigen with accessory cells and cognate helper T cells and/or B lymphocytes in the local micro-environment of the gut and of the lung mucosae, an immune response may ensue, the characteristics of which may be influenced by several factors, including the nature of the antigen, the type of accessory cells and lymphocytes involved, and the genetic background of the host. However, ingestion or inhalation of antigens may also result in the development of a state of peripheral immunological tolerance, a situation characterized by the fact that immune responses in non-mucosal tissues will not develop even if the antigen initially encountered in the digestive tract mucosa or the respiratory mucosa is reintroduced in the organism by a non-mucosal route, such as by parenteral injection. Since this phenomenon is exquisitely specific of the antigen initially ingested or inhaled, and thus does not influence the development of systemic immune responses against other antigens, its use has become an increasingly attractive strategy for preventing and possibly treating illnesses associated or resulting from the development of untoward and/or exaggerated immunological reactions against specific antigens encountered in non-mucosal tissues.
The phenomenon of mucosally induced systemic tolerance may involve all types of immune responses known to be inducible by the systemic introduction of Ag, such as the production of antibodies and the development of cell-mediated immune responses to said Ag. Mucosally induced immunological tolerance has therefore been proposed as a strategy to prevent or to reduce the intensity of allergic reactions to chemical drugs (Chase, M W. 1946. Inhibition of experimental drug allergy by prior feeding of the sensitizing agent. Proc. Soc. Exp. Biol. 61:257-259). It has also been possible to prevent or decrease the intensity of immune reactions to systemically introduced soluble protein antigens and particulate antigens such as red cells in experimental animals and in humans by the oral administration of red cells (Thomas H C and Parrot D M V 1974. The induction of tolerance to soluble protein antigens by oral administration. Immunology 27:631-639; Mattingly, J. and Waksman B. 1978. Immunological suppression after oral administration of antigen. Specific suppressor cells found in rat Peyer's patches after oral administration of sheep erythrocytes and their systemic migration. J. Immunol. 121:1878; Bierme, S. J.; Blank, M.; Abbal, M.; Fournie, A. 1979. Oral Rh treatment for severely immunized mothers. Lancet, 1:605-606).
The phenomenon of mucosally induced systemic tolerance can be utilized to reduce or suppress immune responses not only against foreign antigens but also against self antigens, i.e. components derived from host tissues. It has thus been possible to decrease the intensity of experimentally induced autoimmune diseases in a variety of animal systems by mucosal deposition of auto-antigens onto the intestinal (by feeding) or the respiratory mucosa (by aerosolization or intranasal instillation of antigens). Thus, oral administration of collagen type II (a prominent type of collagen found in joint cartilage) has been shown to suppress or decrease the intensity of experimental autoimmune arthritis, a disease that can be induced in certain strains of rodents by injection of collagen type II together with Freund's complete adjuvant or by injection of Mycobacterium tuberculosis (a component of the former adjuvant) alone (Thompson, H S G and Staines, N A. 1986. Gastric administration of type II collagen delays the onset and severity of collagen-induced arthritis in rats. Clin. Exp. Immunol. 64:581; Nagler-Anderson, C., Bober L A, Robinson, M E, Siskind G W, Thorbecke G J. 1986. Suppression of type II collagen-induced arthritis by intra-gastric administration of soluble type II collagen. Proc. Natl. Acad. Sci. USA 83:7443; Zhang, J Z, Lee, C S Y, Lider, O. and Weiner H L. 1990. Suppression of adjuvant arthritis in Lewis rats by oral administration of type II collagen. J. Immunol. 145:2489-2493). Similarly, it has been possible to suppress an experimental form of autoimmune uveoretinitis by oral administration of S-antigen, a retinal autoantigen that can induce a form of uveoretinitis when injected in animals (Nussenblatt, R B, Caspi, R R, Mahdi R, Chan, C C, Roberge, R., Lider, O., Weiner, H L. 1990. Inhibition of S-antigen induced experimental auto-immune uveoretinitis by oral induction of tolerance with S-antigen. J. Immunol. 144:1689-1695). Experimental autoimmune encephalitis, a chronic relapsing demyelinating disorder that can be induced in certain strains of rodents by injection of purified myelin basic protein or crude spinal cord homogenate together with adjuvant, can be suppressed partially or completely if animals are given MBP or MBP fragments by the oral (feeding) or respiratory (aerosol) route (Bitar D M and Whitacre C C. 1988. Suppression of autoimmune encephalomyelitis by the oral administration of myelin basic protein. Cell Immunol. 112:364; Higgins P J and Weiner H L. 1988. Suppression of experimental autoimmune encephalitis by oral administration of myelin basic protein and its fragments. J. Immunol. 140:440-445; Weiner H L, Al-Sabbagh A and Sobel R. 1990. Antigen driven peripheral immune tolerance: suppression of experimental autoimmune encephalomyelitis (EAE) by aerosol aministration of myelin basic protein. FASEB J (Abstr.) 4(7):2102). Furthermore, oral administration of insulin has been reported to suppress auto-immune immune diabetes in mice (Zhang Z J, Davidson L, Eisenbarth G and Weiner H L. 1991. Suppression of diabetes in non obese diabetic mice by oral administration of porcine insulin. Proc. Natl. Acad. Sci. (USA) 88:10252-10256). More recently, suppression of experimental autoimmune myasthenia gravis has been achieved after oral administration of acetylcholine receptor (Wank Z Y, Qiao J and Link H. 1993. Suppression of experimental autoimmune myasthenia gravis by oral administration of acetylcholine receptor. J. Neuroimmunol. 44:209-214).
It has also been shown that the enteric administration of schistosome eggs in mice could prevent the development or decrease the intensity of hepatic and intestinal granulomatous reactions, which are chronic T cell-mediated inflammatory immune reactions that develop around schistosome eggs during infestation by the parasite Schistosoma (Weinstock J V, Blum A M and Kassab J T. 1985. Induction of granuloma modulation in murine schistosomiasis mansoni by enteric exposure to schistosome eggs. J. Immunol. 135:560-563).
Much in the same way, oral administration of antigens has been proposed to prevent and/or treat allergic reactions to common allergens such as house dust components or substances present in grass pollen (Rebien W, Puttonen E, Maasch H J, Stix E and Wahn U. 1982. Clinical and immunological response to oral and subcutaneous immunotherapy with grass pollen extracts. A prospective study. Eur. J. Pediatry 138:341-344; Wortmann F. 1977. Oral hyposensitization of children with pollinosis or house dust asthma. Allergol et Immunopathol. 5:15-26).
Although the above examples indicate that mucosal administration of foreign as well as self antigens offers a convenient way for. inducing specific immunologic tolerance, the applicability to large scale therapy in human and veterinary medicine remains limited by practical problems.
Indeed, to be clinically broadly applicable, mucosally-induced immunological tolerance must also be effective in patients in whom the disease process has already established itself and/or in whom potentially tissue-damaging immune cells already exist. This is especially important when considering strategies of tolerance induction in patients suffering from or prone to an autoimmune disease, an allergic condition, or a chronic inflammatory reaction to a persistent microorganism. Current protocols of mucosally induced tolerance have had limited success in suppressing the expression of an already established state of systemic immunological sensitization (Hansson D G, Vaz N M, Rawlings L A and Lynch J M. 1979. Inhibition of specific immune responses by feeding protein antigens. II. Effects of prior passive and active immunization. J. Immunol. 122:2261-2266).
Most importantly, and by analogy with mucosal vaccines aimed at inducing immune responses-to infectious pathogens, induction of systemic immunological tolerance by mucosal application of most antigens requires considerable amounts of tolerogen/antigen and, unless the tolerogen/antigen is administered repeatedly over long periods of time is of relatively short duration. A likely explanation is that most antigens are extensively degraded before entering a mucosal tissue and/or are absorbed in insufficient quantities. It has thus been widely assumed that only molecules with known mucosa-binding properties (examples of mucosa-binding molecules are listed in Table I below, see also reviews such as Mirelman D. 1986. Microbial lectins and agglutinins, Properties and biological activity, pp. 84-110, Wiley, New York) can induce local and systemic immune responses when administered by a mucosal route, such as the oral route, without inducing systemic immunological tolerance (de Aizpurua H J and Russell-Jones G J. 1988. Oral vaccination. Identification of classes of proteins that provoke an immune response upon oral. feeding. J. Exp. Med. 167:440-451). A notable example is cholera toxin, one of the most potent mucosal immunogens known so far (Elson C O and Ealding W. 1984. Generalized systemic and mucosal immunity in mice after mucosal stimulation with cholera toxin. J. Immunol. 132:2736) and which when administered simultaneously with an unrelated antigen by the oral route can also prevent induction of systemic immunological tolerance to said antigen (Elson C O and Ealding W. 1984. Cholera toxin did not induce oral tolerance in mice and abrogated oral tolerance to an unrelated antigen. J. Immunol. 133:2892).
Based on these observations, mucosal administration of antigens coupled to mucosa-binding molecules such as cholera toxin or its mucosa-binding fragment cholera toxin B subunit, has been proposed as a strategy to induce local and systemic immune responses rather than systemic tolerance (McKenzie S J and Halsey J F. 1984. Cholera toxin B subunit as a carrier protein to stimulate a mucosal immune response. J. Immunol. 133:1818-1824; Nedrud J G, Liang X, Hague N and Lamm M E. 1987. Combined oral/nasal immunization protects mice from Sendai virus infection. J. Immunol. 139:3484-3492; Czerkinsky C, Russell M W, Lycke N, Lindblad M and Holmgren J. 1989. Oral administration of a streptococcal antigen coupled to cholera toxin B subunit evokes strong antibody responses in salivary glands and extra-mucosal tissues. Infect. Immun. 57:1072-1077; de Aizpurua H J and Russell-Jones G J. 1988. Oral vaccination. Identification of classes of proteins that provoke an immune response upon oral feeding. J. Exp. Med. 167:440-451; Lehner T, Bergmeyer L A, Panagiotidi C, Tao L, Brookes R, Klavinskis L S, Walker P, Walker J, Ward R G et al. 1992. Induction of mucosal and systemic immunity to a recombinant simian immunodeficiency viral protein. Science 258(5036):1365-1369).